- T cell activation is dependent on 3 signals
January 10, 2016
Central Dogma of T cell activation may be too simplistic
Central Dogma of T cell activation may be too simplistic
- The actuality is that the biology of GVHD is likely much more complex
A Mixed Lymphocyte Reaction (MLR) is our best in vitro model of this complexity
- PBMCS from two donors, irradiate one fraction and label other
A Mixed Lymphocyte Reaction (MLR) is our best in vitro model of this complexity
- PBMCS from two donors, irradiate one fraction and label other
- Mix together
A Mixed Lymphocyte Reaction (MLR) is our best in vitro model of this complexity
- PBMCS from two donors, irradiate one fraction and label other
- Mix together
- Allow alloactivation of T cells to proceed over the following 3-5 days
A Mixed Lymphocyte Reaction (MLR) is our best in vitro model of this complexity
- PBMCS from two donors, irradiate one fraction and label other
- Mix together
- Allow alloactivation of T cells to proceed over the following 3-5 days
- Dividing T cells will dilute out the green label (CFSE) and can be quantified flow cytometrically
The MLR through the lens of single-cell RNA-seq
- Dividing T cells will dilute out the green label (CFSE) and can be quantified flow cytometrically
Overview of experiment
- Three donors were utilized (A, B, C)
- MLR was set up using each donor as a responder on each other donor (e.g A on B, A on C, B on A, B on C)
- Controls included T cells cultured with anti-CD3/anti-CD28 microbeads
- Cells were harvested and sorted by CFSE dilution (e.g. Division 0, Division 1, … Division 4)